Brucellae are gram-negative, facultative intracellular bacteria which are one of the most common causes of abortion in animals. In addition, they are the source of a severe zoonosis. In this trial, we evaluated the effect of oral inoculation of Brucella abortus RB51 in mice against a challenge infection with B. abortus 2308. First, we showed that a gastric acid neutralization prior to the oral inoculation contributed to a more homogeneous and consistent infection with both vaccine strain B. abortus RB51 and virulent strain B. abortus 2308. Successively, we assessed the clearance and the immune response following an oral infection with B. abortus RB51. Oral inoculation gave a mild infection which was cleared 42 days after infection, and it induced a delayed humoral and cell-mediated immune response. Finally, we immunized mice by oral inoculation with B. abortus RB51, and we challenged them with the virulent strain B. abortus 2308 by an oral or intraperitoneal route 42 days after vaccination. Oral inoculation of B. abortus RB51 was able to give protection to mice infected with the virulent strain B. abortus 2308 by the oral route but not to mice infected intraperitoneally. Our results indicate that oral inoculation of mice with B. abortus RB51 is able to give a protective immunity against an oral infection with virulent strains, and this protection seems to rely on an immune response at the mucosal level.Brucellae are gram-negative, facultative intracellular bacteria which are one of the most common causes of abortion in animals. In addition, they are the source of a severe zoonosis, which is characterized in human beings by a multitude of somatic complaints, including fever, sweats, anorexia, fatigue, malaise, weight loss, and depression (15). Immune response to Brucella spp. have been studied mainly with mice. In this animal model, protective immunity seems to be mediated by both humoral and cellular immune responses (2). In particular, in vivo studies indicate that an immune response mediated by both CD4 ϩ and CD8 ϩ T lymphocytes is important in controlling infection. Within these subsets, gamma interferon-producing CD4 ϩ T cells play a prominent role in recovery from infection (23). Vaccination against Brucella infections in animals is usually performed by parenteral administration of live attenuated smooth Brucella abortus S19. This vaccine is effective in conferring protection against virulent strains of B. abortus, but it also has a number of disadvantages, including causing abortion in pregnant animals, being pathogenic for humans, and inducing a humoral response which is similar to the response after natural infection, thus hampering procedures for the detection of infected animals by serological testing. In particular, the similarity in humoral responses is of great importance in field conditions because it is impossible, with B. abortus as the antigen, to serologically discriminate between vaccinated and infected animals (11). Recently, a stable, lipopolysaccharide O-antigen-deficient mutant of virulent...